tbuckley/cabot-search huggingface dataset:
Abstract P3068: Hematopoietic Loss Of Y-chromosome In Circulating Monocytes Derived From Heart Failure Patients Associates With Cardiac Fibrotic Signaling
Authors: Wesley T Abplanalp, Sebastian Cremer, David John, Maximilian Merten, Bianca Schuhmacher, Igor Macinkovic, Stefanie Dimmeler, Andreas M Zeiher
Journal: Circulation Research (2023)

Background: Hematopoietic mosaic loss of Y chromosome (LOY) correlates with age and associates with increased cardiovascular disease incidence. Moreover, new murine studies propose that LOY could promote cardiac fibrosis by stimulating cardiac infiltration and activation of circulating macrophages. Yet, specific profiles of hematopoietic LOY cells in patients with heart failure (HF) are missing. Methods and Results: To decipher the genetic profile of human circulating immune cells and assess the effects of LOY, we performed single-cell RNA-sequencing (scRNA-seq) with immune cells of 10 male patients having chronic ischemic HF (mean age 67.3 years, ejection fraction 34.4%). We bioinformatically sorted immune cells by presence or absence of Y-chromosome-associated genes. Relative distribution of LOY was assessed, with monocytes having the greatest absolute count of LOY cells with no significant change in distribution among cell types. Given the abundance of monocytic LOY cells, we examined LOY monocytic cells transcriptional profiles (29810 Y-carrier monocytes, 2205 LOY monocytes) in an unbiased manner. Using a patient-specific, paired analysis, LOY monocytes showed significantly diminished expression of TGF-ß inhibiting genes (SMAD7, TGIF2) in LOY cells versus Y-carrier cells. Opposingly, from 193 upregulated genes, LOY cells showed elevated markers associated with monocyte/macrophage-mediated tissue damage and pro-fibrotic cardiac remodelling (S100A8, CLEC4D, TLR2). Gene ontology terms derived from upregulated genes in LOY cells further supported a mechanistic link to cardiac fibrosis by calling terms associated with TLR signaling, which are known to enhance cardiac fibrosis. Conclusion: This is the first study to study the genetic signature of LOY monocytes in HF patients. Our results considerably extend very recent experimental results in mice that LOY sensitizes macrophages to enhanced signaling associated with cardiac fibrosis. Reduced TGF-ß inhibitory molecule expression and enhanced TLR signaling may contribute to the aggravation of heart failure by LOY in patients with HF.
Abstract P3046: The Effect Of MicroRNA-367-3p And MicroRNA-148a-3p On Vascular Endothelial Cells In Context Of Senescence And Infarct Healing
Authors: Laura Hehl, Katrin Kalies, Fiene Daniel, Kai Knoepp, Jochen Dutzmann, Daniel SEDDING, Jan Michael Köster, Simon Guerlach, Solveig Boettcher
Journal: Circulation Research (2023)

To improve the healing after myocardial infarction (MI), a well-coordinated angiogenic response is necessary. MicroRNAs (miRNAs) were identified as regulators of angiogenesis, however, strategies to improve angiogenesis by targeting miRNAs are challenging. Here, we evaluate miRNA-367-3p and 148a-3p as novel miRNA candidates regarding their therapeutic potential toward neovascularization after MI. Following expression analysis of the respective miRNAs in human vascular endothelial cells (EC) and mice tissue, their contribution to EC function after either pre- or anti-miR transfection was assessed. Further, potential targets were identified and studied on mRNA and protein level. Initial screenings show that miRNA-148a-3p and miRNA-367-3p are differently expressed in both a restenosis model via wire-induced injury and aged mice tissue (p&lt;0.05). Under growth conditions, miRNA-148a-3p expression was upregulated in ECs. In senescent ECs, miRNA-367-3p shows a significant 50-fold upregulation of its expression (p&lt;0.01); miRNA-148a-3p is also overexpressed in senescent ECs (p&lt;0.01). Hypoxia increases the effect observed in senescence (p&lt;0.05). Downregulation of either miRNA-367-3p or miRNA 148a-3p leads to an improvement in migration capacity (p&lt;0.0001) and proliferation of ECs (p&lt;0.0001). Systematic target research in silico identified DKK3 as potential target of miRNA-367-3p and DDX6 as potential target of miRNA-148a-3p. Downregulation of either miRNA-148a-3p or miRNA-367-3p verified these predictions by significantly upregulating DKK3 respectively DDX6. In further experiments, we study the dysfunctional effect of miRNA-148a-3p on vascular endothelial cells by targeting DDX6 as well as the anti-angiogenic impact of miRNA-367-3p by targeting DKK3. Summarizing different expression levels of miRNA-367-3p and miRNA-148a-3p exerted distinct effects on EC function, especially in senescent cells and under hypoxic conditions. Our experimental data regarding cellular function show that these miRNAs are involved in the angiogenetic response and hold the potential to serve as therapeutic targets. In future experiments, we will evaluate the miRNA potential on MI healing in vivo.
Abstract P3179: Tristetraprolin Regulates Metabolic And Epigenetic Remodeling In Cardiomyocytes Post Myocardial Infarction
Authors: Amir Mahmoodzadeh, Ashley E Akrami, Mohammad Keykhaei, Navid Koleini, Hossein Ardehali
Journal: Circulation Research (2023)

Myocardial infarction (MI) triggers profound remodeling in the infarcted, border (BZ), and remote zones of the myocardium. Here, we studied the metabolic remodeling and epigenetic modifications that occur in response to inflammation in the BZ, focusing the role of an mRNA-binding protein that causes degradation of its target mRNA molecules, tristetraprolin (TTP). Inflammatory cells were observed to increase in the BZ, associated with the release of IL-1B and IFN-γ. Among proteins that are altered in the BZ cardiomyocytes, TTP displayed a significant increase in its mRNA and protein levels in the BZ. Isotope tracing using U-13C6 glucose in TTP-overexpressing cells showed that TTP overexpression results in an increase in TCA metabolites upstream of alpha ketoglutarate dehydrogenase (αKGDH), and a decrease of downstream metabolites, suggesting blocked activity at the level of αKGDH. We then identified the mRNA of the E2 component of αKGDH, Dihydrolipoamide S-Succinyltransferase (DLST), to be a target of TTP. Consistent with these findings, DLST mRNA and protein levels are decreased in BZ. DLST needs to be lipoylated to become active, and our data indicate that the mRNA for a key enzyme in DLST lipoylation (lipoic acid synthase, LIAS) is also a target of TTP. Overexpression of TTP led to a decrease in the level of DLST lipoylation. Epigenetic studies showed that, overexpression of TTP resulted in significant decrease in acetylation and methylation of H4K20. Finally, we demonstrated that heart specific TTP KO mice are protected against ischemic injury compared to WT mice, indicating that deletion of TTP alters the remodeling of the BZ, through its role in the regulation of αKGDH and production of αKG in response to infiltration of inflammatory cells.
Abstract P3027: Potasium Adaptation Down-regulates PSGL-1 And I-CAM-1 Gene Expressions In Salt -Induced Hypertensive Sprague Dawley Rats
Authors: Olutayo I Ajayi, MARTHA O OBAYUWANA
Journal: Circulation Research (2023)

Background: There is an increasing prevalence of hypertension and dietary salt intake has been linked as its risk factor. Although many studies have elucidated this association, the mechanisms by which the increase in salt intake leads to development hypertension have been postulated but yet inconclusive. P - Selectin glycoprotein ligand-1(PSGL-1) and Intercellular adhesion molecule (I-CAM-1) have been implicated in the development of salt sensitive hypertension through vascular inflammation and injury. The role of potassium in blood pressure regulation has been severally reported. Scarce reports of the association of potassium adaptation with high blood pressure development form the basis of this study. We aimed at defining the role of potassium adaptation in the development of salt -induced hypertension in Sprague Dawley rats. Methods: This study was divided into two phases: the induction and confirmation of hypertension and the effect of potassium adaptation. The test group received high salt diet (8% NaCl) for 6 weeks while the control animals were fed with standard rat chow and given water ad libitum. Phase II comprised of pre-treated animals with KCl before induction of hypertension and post induction treatment group with KCl. Blood pressures were monitored as well as blood samples taken for analysis of the gene expressions of I-CAM-1 and PSGL-1 using Standard Laboratory methods Results: We observed a spontaneous significant increases in the blood pressures of the test animals as from the 4 th week compared with the control (P&lt;.05, respectively), the potassium -adapted group and KCl post treated groups exhibited relatively stable blood pressure compared with the control. There were significant increases in the expression of PSGL -1 and I-CAM-1 protein in all the test groups but lower in the potassium adapted and post-treated animal groups when compared with hypertensive group. The potassium adapted group did not develop hypertension throughout the period. Conclusion: Raised arterial blood pressure is inducible by high salt diet. This is associated with an increased expressions of PSGL-1 and ICAM-1. Potassium adaptation inhibited the development of hypertension and down-regulate PSGL-1 expression significantly.
Abstract P2098: The Effect Of Cyclic Stretch On Cardiac Fibroblasts
Authors: Fjodor Bekedam, Marisa Smit, rowan Smal, Anton Vonk Noordegraaf, Harm J Bogaard, Marie J Goumans, Frances S De, Aida Llucia-Valldeperas
Journal: Circulation Research (2023)

Introduction: Cardiac fibrosis is the accumulation of extracellular matrix proteins in the interstitium and contributes to heart failure. The main source of cardiac fibrosis is cardiac fibroblasts (cFBs), which are activated as a result of pressure overload. Activated fibroblasts, also known as myofibroblasts, express α-smooth muscle actin (α-SMA) and have an increased production of collagen. Here, the objective was to study the behavior of induced pluripotent stem cell (iPSC) derived cFBs in response to pro-fibrotic cytokines and mechanical stress. Methods: Confluent iPSC monolayers were differentiated into cFBs using the protocol in Figure 1A. Afterwards, the differentiated cells were exposed to 10 ng/ml transforming growth factor β (TGFβ) and 10% cyclic stretch at 1 Hz for 3 days using the Flexcell FX-6000 system. Results: iPSC-cFBs had a fibroblast-like morphology. Furthermore, the presence of cardiac ( GATA4, TCF21 ) and fibroblast ( VIM, PDGFRα, DDR2 ) markers at gene and protein levels confirmed the cFB identity. TGF-β stimulation resulted in increased mRNA and protein levels of α-SMA. Additionally, stretching reduced collagen 1 mRNA and protein expression (Fig. 1B/C). Furthermore, stretching was able to protect the cells from TGF-β stimulation, significantly reducing α-SMA and collagen 1 protein and mRNA levels. This was associated with decreased mRNA expression of TGF-β downstream targets ( PAI1). Conclusion: Cyclic mechanical stretch reduces cFB activation and can protect against TGF-β stimulation, resulting in reduced collagen production and transdifferentiation into myofibroblasts.
Abstract P3121: Overcoming Gender Bias In Cardiovascular Research Using Mice Models By Monitoring Metabolic Differences Between Genders Using Indirect Calorimetry
Authors: Yuka Kimura, Manish K Gupta
Journal: Circulation Research (2023)

Background: Metabolic differences between genders have been observed in humans, with studies reporting differences in body composition, metabolic rates, and response to dietary interventions and how they may affect the risk of cardiovascular disease (CVD) and diabetes. However, most research in CVD models has been conducted exclusively on male mice, resulting in strong gender bias. Moreover, in CVD research investigating differences in hypertension, it is important to note the sexual dimorphisms for these pathologies that result from genetic and hormonal differences. More specifically, age-related CVD rates are lower for women, which has been associated with the protective role of sex steroid hormones in women, which affect the prevalence of metabolic syndrome. However, further research is required to investigate the exact role of these hormones in explaining the lower age-specific CVD rates and the metabolic differences between males and females. Methodology and Result: Adult wild-type C57B6 mice were housed individually in a metabolic cage. We used the indirect calorimetric method to monitor oxygen consumption, carbon dioxide release, respiratory exchange ratio (RER), and energy expenditure of individual mice. Our initial study shows female mice have better volunteer exercise capacity than male mice. Conclusion: This study will address a significant gap in the literature by investigating metabolic differences between genders without gender bias. Our study's findings will provide a foundation for future research in CVD and contribute to developing gender-specific interventions for metabolic disorders.
Abstract P3172: Elamipretide Reduces Mitochondrial Hyperacetylation In Left Ventricular Myocardium Of Dogs With Chronic Heart Failure
Authors: Ramesh C Gupta, Vinita S Gupta, Kristina Castle, Hani N Sabbah
Journal: Circulation Research (2023)

Background: The failing heart manifests abnormalities of mitochondrial (MITO) function characterized by reduced respiration and membrane potential and increased acetylation of MITO proteins. The cause(s) of MITO protein hyperacetylation is unclear, but can be due to reduced sirtuin 3 (Sirt3) possessing deacetylase activity and increased CD38 possessing NADase activity and increased NADH/NAD ratio. Long-term therapy with elamipretide (ELAM), a MITO-targeting tetra-peptide, was shown to improve MITO function in dogs with microembolization-induced heart failure (HF). Hypothesis: We tested the hypothesis that MITO hyperacetylation caused by dysregulation of Sirt3 and CD38 and increased NADH/NAD ratio in MITO of left ventricular (LV) myocardium of HF dogs can be normalized after chronic therapy with ELAM. Methods: MITO fractions were isolated from LV tissue of 14 HF dogs randomized to 3 months monotherapy with subcutaneous injections of ELAM (HF+ELAM, 0.5 mg/kg once daily, n=7) or no therapy at all (control, HF-CON, n=7) and from LV tissue of 6 normal (NL) dogs for comparison. An NAD/NADH quantitation kit was used to determined levels of NADH and NAD in MITO fractions. Protein levels of CD38, Sirt3, and porin, an internal MITO loading control, were determined in Mito-SDS-extracts using Western blotting and band intensity expressed in densitometric units (du). Protein levels of CD38 and Sirt3 were normalized to porin. Results: Protein levels of porin were similar among NL, HF-CON and HF+ELAM dogs. In HF-CON dogs, NADH/NAD ratio increased (0.70 ± 0.14 versus 0.32 ± 0.05, p&lt;0.05 versus HF) compared to NL dogs and decreased significantly in HF+ELAM (0.36 ± 0.02, p&lt;0.05 versus HF-CON). In HF-CON dogs, Sirt3 (0.50 ± 0.0.05 versus 1.24 ± 0.13, p&lt;0.05 versus NL) was significantly reduced and CD38 (1.69 ± 0.12 versus 0.70 ± 0.03, p&lt;0.05 versus NL) was significantly increased compared to NL dogs. Treatment with ELAM normalized Sirt3 (1.0 ± 0.11, p&lt;0.05 vs. HF-CON) and CD38 (0.92 ± 0.051, p&lt;0.05 vs. HF-CON). Conclusions: ELAM normalized levels of Sirt3 and CD38 and reduced NADH/NAD ratio in MITO of LV of HF dogs suggesting normalization of MITO acetylome. The later may be partly responsible for improved MITO function and possibly LV function in HF dogs treated with ELAM.
Abstract P3111: Sex Differences In The Proteomic Response To Chronic Myocardial Ischemia In Swine
Authors: Mark Broadwin, Dwight Harris, Sharif A Sabe, Cynthia Xu, Mohamed Sabra, Ruhul Abid, Frank W Sellke
Journal: Circulation Research (2023)

Intro: There are known sex differences in response to ischemic insult yet the mechanisms behind these differences have not been fully elucidated. Previous work in our lab has demonstrated female sex confers a relative protective effect on CO in the setting of chronic ischemia. The aim of this study was to use total proteomic analysis in order to uncover the mechanisms behind sex specific responses to myocardial ischemia in a porcine model. Methods: Eight 11-week old pigs (3 female, 5 male) underwent surgical placement of an ameroid constrictor to the left circumflex artery via left thoracotomy in order to induce chronic myocardial ischemia. After seven weeks, pigs were euthanized and myocardial tissue was harvested for total proteomic analysis. Results: When compared with non-ischemic male tissue non-ischemic female tissue demonstrated upregulation of two variants of LMOD2 and down regulation of five variants of ACADS (Figure 1A). However, when comparing ischemic female tissue with ischemic male tissue there were 626 upregulated proteins and 194 down regulated proteins (Figure 1B). Enrichment analysis revealed upregulation of certain metabolic pathways including the citric acid cycle and glycolysis/gluconeogenesis (Figure 1C) while there was down regulation in pathways involved in cardiac muscle contraction and dilated cardiomyopathy (Figure 1D). There was no difference in serum glucose, HDL, LDL, or total cholesterol (P &gt; 0.05 for all) (Figure 1E). Conclusion: There were clear sex differences in response to chronic cardiac ischemia. Proteins involved in metabolic pathways and myocyte function appear to be particularly affected.
Abstract P2095: Capillary Stalling by Neutrophils is a Novel Mechanism Underlying Myocardial Hypoperfusion in Heart Failure with Preserved Ejection Fraction
Authors: Anne E Buglione, David M Small, Daniel A Rivera, Nathaniel Allan-Rahill, Soseh Hovasapian, Chris B Schaffer, Nozomi Nishimura
Journal: Circulation Research (2023)

Background: Chronic inflammation and myocardial hypoperfusion are implicated in the pathogenesis of heart failure with preserved ejection fraction (HFpEF), but the cellular and molecular mechanisms are unknown. Intravital cardiac multiphoton microscopy (MPM) enables direct visualization of blood flow and inflammatory cells within the capillary bed of the actively beating heart. We hypothesize that transient arrest of neutrophils in myocardial capillaries leads to perfusion deficits, resulting in hypoxia and driving disease progression. Methods: Male and female 8-12-week-old C57BL/6 mice received a high fat diet and L-NAME in drinking water (HFpEF) or standard diet and water (Chow) for 15 weeks. Intravital imaging of the actively beating heart was performed in mechanically ventilated mice to visualize neutrophils and capillary perfusion. An antibody against the neutrophil surface marker Ly6G (αLy6G) was administered intraperitoneally to test the effect of neutrophil depletion. Results: In vivo MPM revealed increases in arrested neutrophils in myocardial capillaries in HFpEF mice compared to Chows. Myocardial hypoxia, assessed by pimonidazole staining, and diastolic function, assessed as mitral e/e’ using echocardiography, were rescued with both extended neutrophil depletion (2mg/kg every 3d for 4 wk) and acutely (24 hrs after a single dose of 4mg/kg). To further test the immediate effects of αLy6G we implemented multi-exposure laser speckle imaging for perfusion and spectral imaging for tissue and blood oxygenation with electrocardiograph-driven reconstruction. Conclusion: Arrested neutrophils within capillaries appear to compromise myocardial perfusion in HFpEF. Neutrophil depletion to restore blood flow improved myocardial oxygenation and diastolic function rapidly. Effects were sustained with prolonged treatment. Capillary stall perfusion deficits may be a future therapeutic target to slow disease progression and offer rapid symptomatic relief.
Abstract P2114: Chromatin Accessibility Governs Mechanosensitive Gene Expression In Cardiac Fibroblasts
Authors: Jordan Morningstar, Tyler Beck, Cortney Gensemer, Rachel Biggs, Taylor Petrucci, Lilong Guo, Russell A Norris
Journal: Circulation Research (2023)

Objective: We recently demonstrated regionalized myocardial fibrosis in patients undergoing surgical repair for Mitral Valve Prolapse. The location of fibrosis suggests a mechanically driven process wherein mechanical stress alters the phenotype of ventricular fibroblasts to promote an increase in tissue stiffness, eventually causing establishment of a fibrotic scar. We sought to better understand how mechanical stiffness affected cellular programs. Methods: We cultured primary human cardiac fibroblasts derived from the same patient on a mechanically stiff (TC Plastic, ~2GPa), as well as a mechanically soft (2kPa) substrate and then performed immunofluorescence, western blotting, RNA Seq, and ATAC Seq. Following differential analysis of omics data, we performed an integration analysis by filterinig differentially accessible chromatin to only include sites that fell within 10 kB of opened/closed chromatin. Finally, we performed a motif enrichment analysis using the online bioinformatics package Homer. Results: Immunofluorescence and western blotting revealed that cells cultured on a stiff substrate expressed canonical markers of fibroblast activation compared with soft substrates (alpha SMA, COL1A1, P&lt;0.0001). RNA Sequencing revealed differential changes in 1615 genes in fibroblasts (FDR adjusted P&lt;1.6*10 -6 ), several of which were subsequently validated using western blot. ATAC Seq identified 5855 regions across the genome which opened/closed in response to different mechanical environments in fibroblasts (adjusted P&lt;0.05). Integrating these datasets revealed 803 Chromatin changes that fell within 10kb of differentially expressed genes, with AP1 and TEAD family transcription factors predicted to be master regulators of closing and opening chromatin, respectively. Proof-of-concept motif validation conducted on differentially expressed chromatin within intron 12 of the MYH10 gene demonstrated presence of a transcriptional repressor with evidence of JunD as a binding partner for this site. Conclusions: The mechanical environment plays a major role in cellular phenotype and regulates chromatin organization and expression of pro-fibrotic genes through changes in chromatin accessibility.